Compounds or other agents which can chemically alter the DNA of a cell are capable of inducing genetic diseases such as Lesch-Nyhan syndrome, hemophilia, sickle cell anemia, and cystic fibrosis. Compounds or agents which have this potential are known as mutagens. Many mutagens have also been found to have the capability of inducing cancer in test animals or human beings, and are thus also carcinogens. It is clearly desirable, therefore, to have methods for determining the potential mutagenicity of such compounds or agents in a practical, efficient and inexpensive manner.
Bacterial mutation assays have been proposed for this purpose. One of the most commonly employed bacterial assays for mutagenicity is known as the Ames assay and employs a set of Salmonella typhimurium strains which are permeable to a wide range of chemicals and also are partially deficient in DNA repair. Ames, B. N., McCann, J. and Yamasaki, E., Mutation Research, 31, 347-379 (1975). In this system, a chemical's mutagenicity is determined by its ability to revert a set of histidine-requiring mutants of S. typhimurium back to histidine prototrofy through reverse mutation of the original DNA lesion or through a second site mutation.
A more recent assay which employs diploid human lymphoid blastoid cell lines has been developed. This assay depends upon the expression of phenotypic resistance to 6-thioguanine or other purines which serve as substrates for hypoxanthine guaninine phosphoribosyl transferase. This assay is described in U.S. Pat. No. 4,066,510 issued to William G. Thilly.
Although assays of the type described above have proven to be reliable, they do involve extensive experiments of cell cultures at various concentrations of mutagen in order to produce meaningful data. Additionally, they do not predict, with high reliability, the minimum concentration of tested mutagent at which mutagenicity occurs.
Another common biological tool is the diffusion bioassay. In such diffusion bioassays, the reaction of bacteria plated on an agar nutrient medium to an added chemical substance is determined. The chemical diffuses outwardly from some initial spot (e.g., the center of the petri dish), and after a certain incubation period, one or concentric rings around the center can be observed which mark areas in which the bacteria have been affected by the chemical. Such systems have been used in a qualitative manner to determine mutagenicity, and an example of such a test has become known as the Ames spot test. See Ames et al., Ibid. Diffusion bioassays have also been employed to determine the potency of an antibiotic by comparing the radius of a sample of unknown potency to the radius of a standard reference. See Hewitt, W., Microbiological Bioassay, Academic Press, New York, 1977.